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Sphingosine-1-phosphate (S1P) is present in the blood plasma and acts as a pivotal intercellular signal transmitter in the immune system by recruiting lymphocytes from the thymus and secondary lymphoid tissues. The plasma S1P concentration is maintained by the supply of S1P from erythrocytes. Previously, we showed that S1P release from erythrocytes is mediated by an ATP-dependent transporter. In this study, we attempted to establish a rapid and reliable method for measuring the S1P transport activity in erythrocytes by using a fluorescent S1P analog, 7-nitro-2-1,3-benzoxadiazol-4-yl (NBD)-labeled S1P. NBD-S1P was released from erythrocytes in a time-dependent manner. The NBD-S1P release was reduced after exposure to glyburide, which is an inhibitor of the S1P transporter in erythrocytes. Moreover, the release of NBD-S1P and S1P from erythrocytes was competitively inhibited by intracellular S1P and NBD-S1P, respectively. These results showed that the erythrocyte S1P transporter exports NBD-S1P. We optimized the sample-preparation conditions and lipid extraction to increase the sensitivity of the assay. Furthermore, we successfully measured NBD-S1P release without lipid extraction by decreasing the concentration of BSA in the assay buffer to 0.1%. This method will be useful for the high-throughput screening of S1P transporter inhibitors using conventional fluorometers.

A 67-year old male with a dual-chamber pacemaker visited for a regular check-up. An unfamiliar message emerged on the display just after placing the programmer wand. We could recognize that the pacemaker had already been in the safe back-up mode of DDI, and the programmer prompted a re-initialization request. We are so surprised because there was no indication of device malfunction the day before in daily monitoring and a 12-lead electrocardiogram revealed normally working in the DDD mode just before checking the device. The pacemaker was immediately re-programmed to the former setting. This phenomenon has not recurred for 12 months. Keywords: Power-on reset, Electromagnetic interference, Pacemaker, Daily monitoring, Transmission interference